Alcohol dependence (AD) is a serious public health concern characterized by alcohol tolerance, physical dependence, and an inability to control one's alcohol intake. It affects approximately 3.8% of adult Americans each year, and causes substantial economic loss annually. Identification of effective biomarkers for AD is critical for A prevention and treatment. Studies of both cell culture and animal models have demonstrated that small non-coding microRNAs (miRNAs), which regulate the expression of their target genes at the post- transcriptional level, are implicated in chronic alcohol consumption-induced neuroadaptations. Emerging evidence supports the presence of miRNAs in extracellular spaces or body fluids (i.e., saliva, serum, urine, etc), suggesting that the extracellular miRNAs are potential biomarkers for disease diagnosis and prognosis. To date, there is very limited information on miRNA expression changes in human AD subjects, and no study is known to have examined miRNA expression alterations in the saliva of AD subjects. The objective of this application is to identify AD-associated salivary miRNAs and explore the possibility of using these salivary miRNAs (in combination with AD-associated SNPs) to predict AD. The central hypothesis is that chronic alcohol consumption induces altered miRNA expression in various tissues, and that these altered expression levels are reflected in the secretion of miRNAs into body fluids such as saliva. This hypothesis will be tested by pursuing three specific aims: (1) profile salivary miRNA transcriptome changes in AD subjects using next- generation sequencing (NGS) in both African Americans and European Americans; and (2) assess the ability of the differentially expressed miRNAs in predicting the status of AD; and (3) combine AD-associated salivary miRNA and AD-associated SNPs (identified in our recent genome-wide association studies) to improve the prediction of AD. We expect to discover a set of AD-associated salivary miRNAs and SNPs that can serve as AD biomarkers. The approach is innovative because miRNA transcriptome alterations in the saliva of AD subjects will be examined by NGS technology, which offers unprecedented sensitivity and specificity in the detection of gene expression. Moreover, integrating miRNA expression and SNP genotype data is expected to greatly improve the prediction of AD. Our long-term goal is to use the identified AD-associated salivary miRNAs as novel diagnostic and prognostic biomarkers and as potential therapeutic targets for AD and AD- related disorders. The proposed research is significant because the identified salivary miRNAs are expected to have clinical implications in monitoring the development of AD. Additionally, the research strategies developed in this proposed study can be applied to identify extracellular miRNAs that are important for other substance use disorders.